Project Summary Studies have identified thousands of single-nucleotide polymorphisms (SNPs) associated with a range of diseases, but the causal nucleotide changes and mechanisms remain largely unknown. A current bottleneck in the field is that characterization of SNP mechanism greatly lags SNP identification. Analyses suggest that causal variants underlying disease risk are often non-coding SNP (ncSNPs) that function through their effects on gene expression. The primary means by which ncSNPs affect gene expression is by altering binding sites for transcription factor-cofactor (TF-CoF) complexes. Therefore, a major challenge in understanding disease susceptibility and etiology is to characterize the mechanisms by which the thousands of ncSNPs disrupt the binding of TF-CoF complexes to alter gene expression. To address this challenge, we have recently developed CASCADE ? a high-throughput microarray- based method to screen the impact of ncSNPs on the DNA-binding of TF-CoF complexes. In this proposal, we will use our CASCADE approach to study TF-CoF complexes in diverse immune cell states, and study their disruption by disease-associated ncSNPs. To determine the cell state-dependence of our ncSNP annotations and TF-CoF complexes, we will examine results using cell lines as well as primary human cells. To assess CoF specificity for TF classes, and which CoFs are affected by the largest number of ncSNPs (i.e., assay coverage), we will study TF-CoF complexes comprising diverse CoFs and CoF subcomponents. Our goal is to develop CASCADE as a general high-throughput platform to biophysically characterize the regulatory complexes affected by ncSNPs.